We are developing methodology which will lead to the characterization of components necessary and sufficient for transcription of a specific gene as chromatin, in vitro. Isolation of the 2 micron plasmid of yeast as a minichromosome has been carried out. Conditions which apparently lead to specific initiation on the trp 1 yeast gene in pLC544 have been derived; the presence of cytoplasmic factors and exogenous homologous RNA polymerase II are mandatory for correct transcription of restriction endonuclease truncated templates. Other experiments involve characterization of the structure of chromatin during fertilization and early embryogenesis. The sea urchin sperm core particle bears certain similarities to other core particles; however, the presence of variant forms of H2A and H2B leads to marked changes in thermal denaturation of the sperm particle and different accessibilities to cutting by NNAaseI when compared to "typical" core particles from other tissues.